KC-4083

Jurkat-NFAT-Luc2-CD3D-KO-1B1 Cell Line

40624

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Background of Jurkat-NFAT-Luc2-CD3D-KO-1B1 Cell Line

The protein encoded by this gene is part of the T-cell receptor/CD3 complex (TCR/CD3 complex) and is involved in T-cell development and signal transduction. The encoded membrane protein represents the delta subunit of the CD3 complex, and along with four other CD3 subunits, binds either TCR alpha/beta or TCR gamma/delta to form the TCR/CD3 complex on the surface of T-cells. Defects in this gene are a cause of severe combined immunodeficiency autosomal recessive T-cell-negative/B-cell-positive/NK-cell-positive (SCIDBNK). Two transcript variants encoding different isoforms have been found for this gene. Other variants may also exist, but the full-length natures of their transcripts has yet to be defined.

Specifications

Catalog Number	KC-4083
Cell Line Name	Jurkat-NFAT-Luc2-CD3D-KO-1B1 Cell Line
Host Cell Line	Jurkat-NFAT-Luc2
Description	Stable Jurkat-NFAT-Luc2 clone with human CD3D gene knockout, No.1B1
Quantity	One vial of frozen cells (≥2-10⁶/vial)
Stability	Stable in culture over a minimum of 10 passages
Application	Drug screening and biological assays
Freezing Medium	RPMI1640+20% FBS+10% DMSO
Propagation Medium	RPMI1640+10% FBS+300 ug/ml Hygromycin B
Selection Marker	Hygromycin B
Morphology	T lymphocyte
Subculture	Split saturated culture 1:5-1:8 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL
Incubation	37 °C with 5% CO₂
Storage	Liquid nitrogen immediately upon receiving
Doubling Time	Approximately 26 hours
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

Jurkat-NFAT-Luc2-CD3D-KO cell line was generated using the CRISPR method.

Characterization

Figure 1: Characterization of Jurkat-NFAT-Luc2-CD3D-KO-1B1 cell line stable clone using PCR sequencing.

Figure 2: Characterization of Jurkat-NFAT-Luc2-CD3D-KO-1B1 cell line stable clone using RT-PCR sequencing.

Figure 3: Characterization of Jurkat-NFAT-Luc2-CD3D-KO-1B1 cell line stable clone using FACS.

Figure 4: Chareacterization of Jurkat-NFAT-Luc2-CD3-KO clones and Jurkat-NFAT-Luc2 cell line were seeded into the 96-well plate, and treated with OKT3 at a maximum concentration of 20μg/mL for 16 hours, then readout with Bright-Glo system.

Cell Resuscitation

Prewarm culture medium (RPMI1640+10% FBS+300 ug/ml Hygromycin B) in a 37°C water bath.
Thaw the frozen vial in a 37°C water bath for 1-2 minutes.
Transfer the vial into biosafety cabinet, and wipe the surface with 70% ethanol.
Unscrew the top of the vial and transfer the cell suspension gently into a sterile centrifuge tube containing 9.0mL complete culture medium.
Spin at ~ 125 × g for 5-7 minutes at room temperature, and discard the supernatant without disturbing the pellet.
Resuspend cell pellet with the appropriate volume of complete medium and transfer the cell suspension into a T25 culture flask.
Incubate the flask at 37°C, 5% CO₂ incubator.
Split saturated culture 1:5-1:8 every 2-3 days; seed out at about 1-3 ×10⁵ cells/mL.

Cell Freezing

Prepare the freezing medium (70% RPMI1640 + 20% FBS + 10% DMSO) fresh immediately before use.
Keep the freezing medium on ice and label cryovials.
Transfer cells to a sterile, conical centrifuge tube, and count the cells.
Centrifuge the cells at 250×g for 5 minutes at room temperature and carefully aspirate off the medium.
Resuspend the cells at a density of at least 3×10⁶ cells/mL in chilled freezing medium.
Aliquot 1 mL of the cell suspension into each cryovial.
Freeze cells in the CoolCell freezing container overnight in a -80°C freezer.
Transfer vials to liquid nitrogen for long-term storage.

References

https://www.ncbi.nlm.nih.gov/gene/915

Use License Agreement

Research Use Only.

Not for use in diagnostic procedures or therapeutic applications.

Redistribution of the cell line or its derivatives is prohibited without prior written permission from Kyinno Biotechnology.